IgG-type Autoantibody Research Articles

Biphasic Production of Anti-ApoB100 Autoantibodies in Obese Humans and Mice.

Obesity is associated with autoimmunity, a phenomenon considered as harmful. Here we show that obese mice and humans produce IgG-type autoantibodies that specifically recognize apolipoprotein B-100 (ApoB100), its native epitope p210, and the synthetic p210 mimotope pB1. By contrast, antibodies against epitopes p45 and p240, which have been associated with atherosclerosis, were not detected in either the humans or mice. In a longitudinal analysis of high fat diet-fed mice, autoantibody production rose with increasing body weight, then decreased and plateaued at morbid obesity. Likewise, in a cross-sectional analysis of sera from 148 human volunteers spanning a wide BMI range and free of comorbidities, the immunoreactivity increased and then decreased with increasing BMI. Thus, the obesity-related ApoB100-specific natural autoantibodies characteristically showed the same epitope recognition, IgG-type, and biphasic serum levels in humans and mice. We previously reported that a pB1-based vaccine induces similar antibodies and can prevent obesity in mice. Therefore, our present results suggest that autoantibodies directed against native ApoB100 may mitigate obesity, and that the vaccination approach may be effective in humans.

Laryngeal Pemphigoid Evolution and Response to Treatment

This study aimed to present our experience in treating laryngeal pemphigoid (LP) patients, including disease course, treatment and treatment response, and to search for predictors of response to treatment. A retrospective cohort study. The medical records of all patients with LP from March 2013 to August 2020 were reviewed. Potential relationships between disease severity and response to treatment and demographics, diagnosis, extent of laryngeal and extra-laryngeal involvement, comorbidities, immunostaining, and serology profile were explored. Eight patients were included in the study (seven females, one male, mean age 79 years, mean follow-up 22 months). Diagnoses included mucous membrane pemphigoid (MMP, n=5), bullous pemphigoid (BP, n=3). Two patients achieved complete laryngeal remission, four achieved partial remission, and two had no remission. The time to achieve laryngeal disease control was longer than for extra-laryngeal disease (P = 0.02). Potential associations were found between the absence of immunoglobulin G (IgG)-type auto-antibodies deposits in the basement membrane zone and a laryngeal disease that responded to topical corticosteroids and between the presence of BP180-C-terminal IgG auto-antibodies and a resistant rapidly progressive laryngeal disease. LP has a spectrum of severity, variable response and is more resistant to treatment. The absence of IgG-type auto-antibodies may indicate a response to topical corticosteroids. Based on our limited observation, the presence of IgG-type auto-antibodies that target BP180-C-terminal domain may indicate a more severe scarring disease. Early recognition of these "high-risk patients" will allow early initiation of advanced systemic treatment that may prevent the irreversible effects of scarring.

Anti-Tyro3 IgG Associates with Disease Activity and Reduces Efferocytosis of Macrophages in New-Onset Systemic Lupus Erythematosus.

Background Systemic lupus erythematosus (SLE) is a disease characterized by the production of a large number of autoantibodies. Defected phagocytosis of macrophage plays an important role in innate immunity in the pathogenesis of SLE. Tyro3 is a receptor responsible for the recognition of apoptotic cells during efferocytosis by macrophages. To investigate the role of Tyro3 receptor in macrophages' efferocytosis of apoptotic cells in SLE, we aimed to reveal the clinical relevance and impact of Tyro3 autoantibody on SLE. Methods The serum levels of IgG-type autoantibody against Tyro3 receptor were detected in new-onset, treatment-naïve SLE patients (n = 70), rheumatoid arthritis (RA) (n = 24), primary Sjögren's Syndrome (pSS) (n = 21), and healthy controls (HCs) (n = 70) using enzyme-linked immunosorbent assay (ELISA). The effects of purified Tyro3 autoantibody from SLE patients on the efferocytosis of human monocyte-derived macrophages were measured by flow cytometry and immunofluorescence. Results The serum levels of IgG-type autoantibody against Tyro3 receptor were significantly elevated in patients with SLE compared to RA, pSS, and HCs (all p < 0.0001). The levels of anti-Tyro3 IgG were positively associated with the SLE disease activity index (SLEDAI) score (r = 0.254, p = 0.034), erythrocyte sedimentation rate (ESR) (r = 0.430, p < 0.001), C-reactive protein (CRP) (r = 0.246, p = 0.049), and immunoglobulin G (IgG) (r = 0.408, p = 0.001) and negatively associated with haemoglobin (Hb) (r = −0.294, p = 0.014). ROC curves illustrated that the anti-Tyro3 antibody could differentiate patients with SLE from HCs. Furthermore, flow cytometry and immunofluorescence demonstrated that purified anti-Tyro3 IgG inhibited the efferocytosis of macrophages (p = 0.004 and 0.044, respectively) compared with unconjugated human IgG. Conclusions These observations indicated that autoantibody against Tyro3 was associated with disease activity and could impair efferocytosis of macrophages. It might be a potential novel disease biomarker and might be involved in the pathogenesis of SLE.

High-resolution epitope mapping by HX MS reveals the pathogenic mechanism and a possible therapy for autoimmune TTP syndrome

Acquired thrombotic thrombocytopenic purpura (TTP), a thrombotic disorder that is fatal in almost all cases if not treated promptly, is primarily caused by IgG-type autoantibodies that inhibit the ability of the ADAMTS13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13) metalloprotease to cleave von Willebrand factor (VWF). Because the mechanism of autoantibody-mediated inhibition of ADAMTS13 activity is not known, the only effective therapy so far is repeated whole-body plasma exchange. We used hydrogen-deuterium exchange mass spectrometry (HX MS) to determine the ADAMTS13 binding epitope for three representative human monoclonal autoantibodies, isolated from TTP patients by phage display as tethered single-chain fragments of the variable regions (scFvs). All three scFvs bind the same conformationally discontinuous epitopic region on five small solvent-exposed loops in the spacer domain of ADAMTS13. The same epitopic region is also bound by most polyclonal IgG autoantibodies in 23 TTP patients that we tested. The ability of ADAMTS13 to proteolyze VWF is impaired by the binding of autoantibodies at the epitopic loops in the spacer domain, by the deletion of individual epitopic loops, and by some local mutations. Structural considerations and HX MS results rule out any disruptive structure change effect in the distant ADAMTS13 metalloprotease domain. Instead, it appears that the same ADAMTS13 loop segments that bind the autoantibodies are also responsible for correct binding to the VWF substrate. If so, the autoantibodies must prevent VWF proteolysis simply by physically blocking normal ADAMTS13 to VWF interaction. These results point to the mechanism for autoantibody action and an avenue for therapeutic intervention.

Celiac disease in children with atopic dermatitis.

Celiac disease (CD) is an autoimmune disorder of the small intestine with highly variable clinical presentation and frequently associated with various immune-mediated diseases. Among these immune-mediated diseases, atopy has been found frequently in individuals with CD. We aimed to study the prevalence of CD in Estonian children with atopic dermatitis (AD), a common multifactorial chronic inflammatory skin disease. We recruited 351 consecutive children with active AD (mean age 5.8yrs, 57.6% boys) at Tallinn Children's Hospital, Estonia. Sera of all patients were tested for total serum immunoglobulin (Ig) A, for IgA- and IgG-type autoantibodies to tissue transglutaminase (IgA-anti-TG2, IgG-anti-TG2) and to deamidated gliadin peptides (IgA-anti-DGP, IgG-anti-DGP). The diagnosis of CD was confirmed histologically by small intestine biopsy according to the European Society of Paediatric Gastroenterology, Hepatology and Nutrition diagnostic criteria. IgA deficiency was detected in nine patients with AD (2.6%), none of whom had IgG-anti-TG2 or IgG-anti-DGP seropositivity. IgA-anti-TG2 positivity was found in 4 (1.1%), IgG-anti-TG2 positivity in 2 (0.6%), IgA-anti-DGP positivity in 11 (3.1%), and IgG-anti-DGP in 10 (2.8%) patients. Celiac disease was confirmed in five (1.4%) patients with AD (95% confidence interval 0.46, 3.32) and all were histologically characterized as Marsh IIIa-IIIc stages and two presented with silent CD. In AD patients, CD prevalence was more than four times as high as in previously studied randomly selected schoolchildren in Estonia. Two patients with AD diagnosed with CD had no symptoms indicative of CD, in spite of extensive histologic changes in the small intestine mucosa. Therefore our study emphasizes the need for evaluating the cost-effectiveness of screening individuals with AD for CD in time to prevent long-term complications.

Increased Levels of Antigen-Bound β-Amyloid Autoantibodies in Serum and Cerebrospinal Fluid of Alzheimer’s Disease Patients

Recent studies have suggested a protective role of physiological β-amyloid autoantibodies (Aβ-autoantibodies) in Alzheimer’s disease (AD). However, the determination of both free and dissociated Aβ-autoantibodies in serum hitherto has yielded inconsistent results regarding their function and possible biomarker value. Here we report the application of a new sandwich enzyme-linked immunosorbent assay (ELISA) for the determination of antigen-bound Aβ-autoantibodies (intact Aβ-IgG immune complexes) in serum and cerebrospinal fluid (CSF) of a total number of 112 AD patients and age- and gender-matched control subjects. Both serum and CSF levels of Aβ-IgG immune complexes were found to be significantly higher in AD patients compared to control subjects. Moreover, the levels of Aβ-IgG complexes were negatively correlated with the cognitive status across the groups, increasing with declining cognitive test performance of the subjects. Our results suggest a contribution of IgG-type autoantibodies to Aβ clearance in vivo and an increased immune response in AD, which may be associated with deficient Aβ-IgG removal. These findings may contribute to elucidating the role of Aβ-autoantibodies in AD pathophysiology and their potential application in AD diagnosis.

IgM-type GM-CSF autoantibody is etiologically a bystander but associated with IgG-type autoantibody production in autoimmune pulmonary alveolar proteinosis

The granulocyte-macrophage colony-stimulating factor (GM-CSF) autoantibody (GMAb) is the causative agent underlying autoimmune pulmonary alveolar proteinosis (aPAP). It consists primarily of the IgG isotype. At present, information on other isotypes of the autoantibody is limited. We detected serum the IgM isotype of GMAb (IgM-GMAb) in more than 80% of patients with aPAP and 22% of healthy subjects, suggesting that a continuous antigen pressure may be present in most patients. Levels of the IgM isotype were weakly correlated with IgG-GMAb levels but not IgA-GMAb, suggesting that its production may be associated with that of IgG-GMAb. The mean binding avidity to GM-CSF of the IgM isotype was 100-fold lower than the IgG-GMAb isotype, whereas the IC(50) value for neutralizing capacity was 20,000-fold higher than that of IgG-GMAb, indicating that IgM-GMAb is only a very weak neutralizer of GM-CSF. In bronchoalveolar lavage fluid from nine patients, IgG-GMAb was consistently detected, but IgM-GMAb was under the detection limit in most patients, confirming that IgM-GMAb is functionally a bystander in the pathogenesis of aPAP. It rather may be involved in the mechanism for development of IgG-GMAb in vivo.

Chemotherapy-resistant intravascular lymphoma accompanied by ADAMTS13 inhibitor successfully treated with rituximab

Intravascular large B-cell lymphoma (IVL) is a subtype of diffuse large B-cell lymphoma (DLBCL), in which lymphoma cells infiltrate blood vessels without mass formation, and its prognosis is generally poor due to diagnostic difficulties. The authors previously described a patient with IVL accompanied by features of thrombotic thrombocytopenic purpura (TTP), expression of ADAMTS13 inhibitor, and a severe deficiency in ADAMTS13 activity [1]. Although standard CHOP chemotherapy was initially effective, the disease gradually became resistant despite intensive high-dose chemotherapy with autologous peripheral blood stem cell support. However, the chimeric anti-CD20 monoclonal antibody, rituximab, induced complete remission (CR) for a prolonged period. We describe the clinical course of this patient and the unique effects of treatment with rituximab alone. A 54-year-old man was admitted to our hospital with the onset of a psychological disorder and convulsions. Laboratory data revealed obvious thrombocytopenia (3.6 9 10/ lL) accompanied by elevated levels of serum lactic dehydrogenase (LDH) and soluble interleukin-2 receptor. Multiple cerebral venous embolisms, bilateral adrenal gland swelling, and hepatosplenomegaly were present without nodal disease. Despite normal chest X-rays and thoracic CT scans, the patient developed rapidly progressive respiratory failure. A definitive diagnosis of IVL was made from transbronchial lung biopsy (TBLB) specimens. Bone marrow smears revealed no obvious lymphoma cell invasion and megakaryocyte hyperplasia without hemophagocytic syndrome (HPS), although the peripheral blood platelet count was markedly decreased. We concluded that the thrombocytopenia was caused by the same pathogenesis as TTP, since plasma ADAMTS13 activity had decreased to below 3%, and active ADAMTS13 inhibitor (IgG-type autoantibody) was confirmed. After standard CHOP chemotherapy cycles, the platelet counts immediately recovered to within normal limits and all symptoms improved. The ADAMTS13 activity improved to 49% and inhibitor levels, which had never been negative, decreased. Whole brain irradiation after chemotherapy resulted in relapse of systemic disease. Although we performed highdose conditioning chemotherapy followed by autologous peripheral stem cell transplantation, respiratory failure and high-grade fever appeared with severe thrombocytopenia and massive serum LDH elevation at day 50 after transplantation. Salvage combination chemotherapies had extremely limited effects against the disease, and we started to administer 375 mg/m of rituximab after its use for CD20-positive DLBCL was permitted according to the regulations established by the Japanese National Health Insurance system. The patient did not develop any major M. Kanno (&) Tumor Center, Nara Medical University Hospital, 840 Shijo-cho, Kashihara 634-8522, Japan e-mail: mkanno@naramed-u.ac.jp

Comprehensive analysis of ADAMTS13 in patients with liver cirrhosis

Decreased plasma ADAMTS13 activity (ADAMTS13:AC) results in the accumulation of unusually large von Willebrand factor multimer (UL-VWFM) and the formation of platelet thrombi. It remains controversial whether or not plasma ADAMTS13:AC decreases in patients with liver cirrhosis (LC), and its relationship to clinical features has not been fully investigated. We measured ADAMTS13:AC and its related parameters in plasma in 33 patients with chronic hepatitis (CH) and in 109 patients with LC. ADAMTS13:AC decreased with increasing severity of liver disease (controls means 100%, CH 87%, Child A-LC 79%, Child B-LC 63%, and Child C-LC 31%), and showed severe deficiency (<3% of controls) in five end-stage LC. Activities measured by act-ELISA strongly correlated with those determined by the VWFM assay and ADAMTS13 antigen. Multivariate analysis showed Child-Pugh score and spleen volume independent factors contributing to ADAMTS13:AC. VWFM patterns were normal in 53% of cases, degraded in 31%, and unusually large in 16%. Patients with unusually large VWFM had the lowest ADAMTS13:AC as well as the highest Child-Pugh score, serum creatinine and blood ammonia levels. Plasma inhibitor against ADAMTS13 detected in 83% of patients with severe to moderate ADAMTS13:AC deficiency mostly showed marginal zone between 0.5 and 1.0 BU/ml. The IgG-type autoantibodies specific to plasma derived-ADAMTS13 was detected by Western blot in only five end-stage LC with severe ADAMTS13:AC deficiency. In conclusion, both plasma ADAMTS13 activity and antigen levels decreased with increasing severity of cirrhosis. An imbalance between the decreased ADAMTS13:AC and its increased substrate may reflect the predisposing state for platelet thrombi formation in patients with advanced LC.

Anti-CD69 autoantibodies cross-react with low density lipoprotein receptor-related protein 2 in systemic autoimmune diseases.

We investigated whether autoantibodies to CD69, one of the earliest markers of lymphocyte activation, exist in the sera of patients with systemic autoimmune disease. Serum samples were obtained from patients with rheumatoid arthritis (RA), systemic lupus erythematosus, and Behcet's disease, and were tested for the presence of anti-CD69 autoantibodies by ELISA and Western blotting using rCD69 fusion proteins. IgG-type autoantibodies to CD69 were detected in the sera of 38.3% of the RA patients, 14.5% of the systemic lupus erythematosus patients, and 4.0% of the patients with Behcet's disease. Among those with RA, the anti-CD69 autoantibody-positive patients had a higher serum level of rheumatoid factors and a more accelerated erythrocyte sedimentation rate than the anti-CD69 autoantibody-negative patients. Further, the predominant epitope on the CD69 molecule to which most of the anti-CD69 autoantibody-positive serum samples exclusively reacted, was mapped at the C terminus of CD69. Of interest, this epitope is homologous to a stretch of amino acids in the protein sequence of low-density lipoprotein receptor-related protein 2 (LRP2), which is a receptor for multiple ligands including beta-very low density lipoprotein and is also an autoantigen responsible for Heymann nephritis in rats. The anti-CD69 autoantibody cross-reacted to LRP2 through the homologous amino acid sequence. To our knowledge, this is the first evidence of the existence of anti-CD69 autoantibodies. This autoantibody may modulate the function of CD69- and LRP2-expressing cells.

Loss of TGF-β signaling contributes to autoimmune pancreatitis

Recent observations suggest that immune response is involved in the development of pancreatitis. However, the exact pathogenesis underlying this immune-mediated response is still under debate. TGF-beta has been known to be an important regulating factor in maintaining immune homeostasis. To determine the role of TGF-beta in the initiation or progression of pancreatitis, TGF-beta signaling was inactivated in mouse pancreata by overexpressing a dominant-negative mutant form of TGF-beta type II receptor in the pancreas, under control of the pS2 mouse trefoil peptide promoter. Transgenic mice showed marked increases in MHC class II molecules and matrix metalloproteinase expression in pancreatic acinar cells. These mice also showed increased susceptibility to cerulein-induced pancreatitis. This pancreatitis was characterized by severe pancreatic edema, inflammatory cell infiltration, T- and B-cell hyperactivation, IgG-type autoantibodies against pancreatic acinar cells, and IgM-type autoantibodies against pancreatic ductal epithelial cells. Therefore, TGF-beta signaling seems to be essential either in maintaining the normal immune homeostasis and suppressing autoimmunity or in preserving the integrity of pancreatic acinar cells.

Treatment of Lupus in NZB/W F1 Mice with Monoclonal Antibody Against Fas Ligand

Since Fas ligand (FasL) can induce apoptosis of Fas-bearing cells, Fas/FasL interactions can play a critical role in maintaining self-tolerance. Fas/FasL interactions in lupus-like autoimmune disease have been well characterized in studies using either Fas or FasL mutant mice. However, the effect of the defective FasL-mediated signaling on the establishment of lupus in other mouse strains, such as NZB/W (B/W) F1, remains uncertain. In the present study, we examined the effect of anti-FasL monoclonal antibody (mAb) on the development of lupus. Treatment of B/W F1 mice with anti-FasL mAb augmented IgG1- and IgG2a-type anti-dsDNA Ab production. However, treatment of B/W F1 mice with anti-FasL mAb also significantly prevented the development of lupus nephritis. These results indicate that Fas/FasL interactions not only regulate IgG-type autoantibody production, but also influence the development of lupus nephritis in B/W F1 mice.

POSTOPERATIVE GIANT RETROPERITONEAL HEMATOMA DUE TO THE APPEARANCE OFCOAGULATION FACTOR VIII INHIBITOR-A CASE REPORT-

A case of giant retroperitoneal hematoma due to the appearance of coagulation factor VIII inhibitor is reported. A 69-year-old man was admitted to the hospital because of hematemesis, and was operated on with a diagnosis of early cancer of the cardiac part of the stomach. After the operation, hemorrhage from ulcer at the anastomosed site occurred which demanded reoperation. Thereafter the patient had hematemesis again but was conservatively managed. Two months later, however, a giant hematoma was abruptly formed at the retroperitoneum and shock occurred. On a coagulation test, a prolongation in APTT alone was noted. Exploration revealed that it was bleeding tendency due to the appearance of the factor VIII inhibitor. The cause was obscure, but the APTT level was normalized for a time after starting the prednisolone regimen. An immunological analysis of the inhibitor demonstrated that it was IgG type autoantibody and IgG1 and IgG4 as IgG subclass. Bleeding tendency due to the appearance of coagulation factor VIII inhibitor often causes severe hemorrhagic symptoms like in this case, with a high mortality rate due to hemorrhage. So the recognition and rapid treatment of the disease are of importance.

Immunological Analysis of Acquired Factor VIII Inhibitor in a Case with Immunologic Disorder

Coagulation factor VIII inhibitor arising in a patient with autoimmune disease was immunologically analyzed. A 63-year-old man who had been diagnosed as suffering from polyarteritis nodosa was treated with prednisolone for 10 years. Severe bleeding tendency developed and coagulation studies demonstrated a high titer of inhibitor to factor VIII:C. As a result of immunological analysis, the inhibitor was found to be IgG type autoantibody having both kappa and lambda light chains. The subclasses were IgG1 and IgG4. The inhibitor recognized the COOH-terminal light chain (72-kDa thrombin fragment) on the factor VIII molecule as an epitope.